Cross talk between oncogene and ant-oncogene via the Crk oncogene product

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Crk oncogene product activates Ras and Rap1. Ras is a well-known oncogene, whereas Rap1 antagonize Ras-dependent oncogenesis. To understand the mechanism by which Crk activates both oncogene and anti-oncogene, we developed a FRET-based single-molecule probe that can monitor the activation of Crk in a living cell. Using this probe, we visualized rapid Crk activation in an EGF-stimulated cell. However, with the prototype probe, we could not determine where in the cell the activation of Crk starts, due to rapid diffusion of the probe. To overcome this flaw, we fused CAAX box of Ki-Ras protein to the probe, expecting the anchoring of probe to the plasma membrane. Indeed, by the use of this CAAX-fused probe, we found that Crk was activated from the peripheral region of the cell. Furthermore, we generated a recombinant adenovirus for the expression of the probe, which enabled us rapid and efficient expression of the probe in a variety of cells. Thus, the development of new probe for the Crk activity visualized the spatio temporal regulation of Crk oncogene products in living cells and promised us for the understanding of the role of Crk oncogene products in the regulation of cell growth and transformation.